Willem F. Decraemer

Refractive index of tissue measured with confocal microscopy

Refractive index of tissue is an essential parameter in many bio-optical experiments, yet little data can be found in literature. Several methods have been proposed to measure refractive index in tissue samples, but all have specific limitations, such as low accuracy, the need for large amounts of tissue, or the complexity of the measurement setup. We propose a new method using a standard confocal microscope and requiring only small tissue samples. A thin slice of tissue is put next to a layer of immersion fluid of exactly the same thickness. The actual thickness of the fluid layer is directly measured with the microscope, as there is no refractive index mismatch. A difference between index of refraction of the tissue and of the immersion medium causes an axial scaling factor. The optical thickness of the specimen is thus measured with the microscope, and as its actual thickness equals the known thickness of the fluid layer, the axial scaling factor is readily determined. From this factor, we calculate the refractive index of the tissue. We use a diffraction model to take the point spread function (PSF) of the microscope into account, so we can determine the index of refraction to a very high accuracy. We demonstrate the method on bovine muscle tissue and find a value of n=1.382+/-0.004, at 592 nm.

On the damped frequency response of a finite‐element model of the cat eardrum

This article presents frequency responses calculated using a three-dimensional finite-element model of the cat eardrum that includes damping. The damping is represented by both mass-proportional and stiffness-proportional terms. With light damping, the frequency responses of points on the eardrum away from the manubrium display numerous narrow minima and maxima, the frequencies and amplitudes of which are different for different positions on the eardrum. The frequency response on the manubrium is smoother than that on the eardrum away from the manubrium. Increasing the degree of damping smooths the frequency responses both on the manubrium and on the eardrum away from the manubrium. The overall displacement magnitudes are not significantly reduced even when the damping is heavy enough to smooth out all but the largest variations. Experimentally observed frequency responses of the cat eardrum are presented for comparison with the model results.

On the degree of rigidity of the manubrium in a finite-element model of the cat eardrum

It has always been assumed that the manubrium is in effect perfectly rigid. In this paper, a more realistic model of the manubrium is incorporated into an existing finite-element model of the cat eardrum. The manubrial thickness is based on a three-dimensional reconstruction from serial histological sections. After a review of the literature, a value of 2 x 10(11) dyn cm-2 is adopted for the Youngs modulus of the bone. The mode of vibration of the model is investigated for different manubrial-thickness values and it is found that a significant degree of manubrial bending occurs in the model for realistic values of manubrial thickness. As a result of the bending, the frequency response at the umbo at high frequencies displays much higher amplitudes and larger phase lags than when the manubrium is rigid. The bending will also affect the displacements transmitted to the ossicular load, and introduce significant errors into estimates of such displacements based on measurements of umbo displacement even at frequencies as low as a few kHz. Recent measurements of manubrium vibrations in the cat ear provide experimental evidence of bending.

Volume displacement of the gerbil eardrum pars flaccida as a function of middle ear pressure

The pars flaccida (PF) is a small region of the eardrum, with elasticity and histology completely different from the rest of the membrane, which has often been attributed a pressure regulating function for the middle ear (ME). In this paper, the volume displacement of the PF as a function of ME pressure is discussed. The deformation of the PF was measured in vitro in five Mongolian gerbil ears, by means of an opto-electronic moiré interferometer. Volume displacement was determined at small intervals in three sequential pressure cycles, in the range of +/- 0.4 kPa, +/- 2 kPa, and again +/- 0.4 kPa. The displacement was found to be a highly non-linear function of pressure, with a strong increase up to 0.4 kPa ME over- or underpressure and remaining nearly unchanged for pressures beyond 0.4 kPa. In all animals, maximal volume displacement was less than 0.5 microl, or 0.2% of total ME air volume. Clear hysteresis was found between the deformations at the same pressure level in the increasing and decreasing parts of the pressure cycles. Membrane behavior in the first 0.4 kPa pressure cycle was significantly different from that in the second 0.4 kPa cycle, which followed the 2 kPa pressure cycle. The results indicate that the ME pressure change regulation function of the PF is limited to very small pressure changes of a few hundred Pa around ambient pressure, and that larger ME pressures cause at least short-term changes in the membranes behavior.

Interferometric measurement of the amplitude and phase of tympanic membrane vibrations in cat.

The amplitude and phase of the tympanic membrane and malleus vibrations were measured over a wide frequency range with a homodyne interferometer. When sound pressure was maintained constant near the tympanic membrane, the malleus frequency response followed the typical pattern up to 10 kHz as measured by previous investigators. At higher frequencies the response changes dramatically. Instead of decreasing with frequency, between 10 and 20 kHz the vibration amplitude oscillates around a value which is only about 20 dB lower than the low frequency plateau level. Measurements of malleus vibration at several points along its length indicate that its mode of vibration changes at high frequencies, and no longer consists of a simple rotational component. All points on the tympanic membrane vibrate in phase with the malleus up to a frequency of 1 kHz. Above 5 kHz discrete resonances are observed, and the response varies strongly with position on the tympanic membrane.

Phase shift method based on object translation for full field automatic 3-D surface reconstruction from moire topograms

A new method fur full field automatic 3-D surface reconstruction is proposed which makes use of multiple contourograms shifted in phase by object translation. The method is demonstrated for shadow moire topography. It is shown that surface reconstruction can be done fast and with a resolution at least 10 times higher than the fringe distance of the measuring setup. Convexity and concavity of the surface are automatically determined. Also shown is the possibility of measuring irregular surfaces with very sudden height jumps.

A method for determining three-dimensional vibration in the ear

In the classical concept of the middle ear function the malleus rotates around a fixed axis which implies that at small amplitudes of vibration its displacement is essentially one dimensional. As a consequence malleus vibrations have been measured previously along a single viewing axis. As a first step in the study of the complete malleus motion we determined the three dimensional components at a single point (umbo) of the manubrium. To define 3-D motion it is in principle necessary to measure the vibrations from widely different observation angles. The viewing angles are limited however in our case by the ear canal geometry to about +/-15 degrees. In order to resolve the 3-D components under these conditions it is necessary to measure the vibration components with high accuracy. Amplitude and phase of the umbo vibrations were measured with a heterodyne interferometer over a wide frequency range (100 Hz to 20 kHz). The system included a two axis goniometer with the axes of rotation positioned at the focal plane of the interferometer objective lens. It was therefore possible to change the viewing angle in small increments around two orthogonal axes while keeping the same point in focus. From a redundant set of measurements the three orthogonal components of vibration were calculated by least squares fitting. The vector sum of the three components gives the three dimensional motion of the observed point. The vibration of the point on the umbo was found not to follow a straight line but an elliptical path instead. The shape of the ellipse and the inclination of the plane of the ellipse with respect to the stationary malleus position changed with frequency. These observations are consistent with our earlier findings that the mode of malleus vibration changes with frequency [Decraemer et al. (1991) Hear. Res. 54, 305-318].

Three-Dimensional Modelling of the Middle-Ear Ossicular Chain Using a Commercial High-Resolution X-Ray CT Scanner

The quantitative measurement of the three-dimensional (3-D) anatomy of the ear is of great importance in the making of teaching models and the design of mathematical models of parts of the ear, and also for the interpretation and presentation of experimental results. This article describes how we used virtual sections from a commercial high-resolution X-ray computed tomography (CT) scanner to make realistic 3-D anatomical models for various applications in our middle-ear research. The important problem of registration of the 3-D model within the experimental reference frame is discussed. The commercial X-ray CT apparatus is also compared with X-ray CT using synchrotron radiation, with magnetic resonance microscopy, with fluorescence optical sectioning, and with physical (histological) serial sections.

Human tympanic membrane deformation under static pressure

The effect of static pressures in the range of plus and minus 1.6 kPa on the shape of tympanic membrane is measured using a non-contacting optical technique on a fresh human temporal bone. Full field data of the deformation are presented as well as cross-sections along two major directions. Strong asymmetry between medial and lateral movements is demonstrated. The displacement of the umbo is compared to other work. The rotation angle of the manubrium in function of pressure is calculated and also compared to other work. It is demonstrated that the rotation angels can not account for the measured movement of the umbo, which leads to the conclusion that for static high pressure levels the classical hypothesis of rotation around a fixed axis has to be abandoned. The comparison with data of TM displacement under dynamic stimuli is discussed.

On the Coupling Between the Incus and the Stapes in the Cat

The connection between the long process and the lenticular process of the incus is extremely fine, so much so that some authors have treated the lenticular process as a separate bone. We review descriptions of the lenticular process that have appeared in the literature, and present some new histological observations. We discuss the dimensions and composition of the lenticular process and of the incudostapedial joint, and present estimates of the material properties for the bone, cartilage, and ligament of which they are composed. We present a preliminary finite-element model which includes the lenticular plate, the bony pedicle connecting the lenticular plate to the long process, the head of the stapes, and the incudostapedial joint. The model has a much simplified geometry. We present simulation results for ranges of values for the material properties. We then present simulation results for this model when it is incorporated into an overall model of the middle ear of the cat. For the geometries and material properties used here, the bony pedicle is found to contribute significant flexibility to the coupling between the incus and the stapes.